BOG (RBBP-9) is a novel cellular protein which interacts with pRb, p107 and p130 through the LXCXE motif. Substitution of a leucine with glutamine in the LXCXE sequence (BOG D) is sufficient to disrupt the interaction between BOG and pRb/p107/p130. Here we show by employing subcellular fractionation, immunohistochemistry and BOG-EGFP1 fusion constructs, that the BOG protein was primarily located in the nucleus in a variety of cell lines. To examine whether the nuclear localization of the BOG protein is dependent on its pRb binding motif, we expressed the fusion proteins BOG-EGFP and BOG D-EGFP, in several mouse, rat and human cell lines. In Saos-2 cells, which contain a functionally inactive pRb, and in pRb -/- mouse embryo fibroblasts, the fusion protein BOG-EGFP was still localized to the nucleus. However, unlike the wild-type, BOG D-EGFP failed to accumulate in the nuclei, and demonstrated a greater cytoplasmic localization. These results indicate that LXCXE motif is critical for both binding of BOG to members of the Rb family and the nuclear localization of the protein, suggesting that the binding with the pRb related proteins p107 and/or p130 may be sufficient to support the nuclear accumulation of the protein in the absence of pRb.